Glucose Metabolism
Pratt and Cornely, Chapter 13
Glycolysis Expectations
• Memorize/learn Figure 13.2• Know overall reaction and stages• Explain chemical logic of each step• Enzyme mechanisms presented in book
Glycolysis
• Ten enzymes that take glucose to pyruvate
• Cytosol• ATP and NADH
Reactions and Enzymes of Glycolysis
• Hexose and triose phases
• Energy input and payoff phases
ATP ATP
ADP ADP2x
Pi + NAD+
NADH
2x
ADP ADP
ATP ATP
2x 2x
Energy Input
Energy Payoff
Know...
• Substrates • Co-substrates • Products • Enzyme names
1. Hexokinase
• Previous concepts: Induced fit, kinase• Energy use/production? • Chemical logic?
Problem 3• (Notice miswording) The DGo’ value for
hexokinase is -16.7 kJ/mol, and the DG value under cellular conditions is similar.– What is the ratio of G-6-P to glucose under standard
conditions at equilibrium if the ratio of ATP:ADP is 10:1?
– How high would the ratio of G-6-P to glucose have to be to reverse the hexokinase reaction by mass action?
2. Phosphoglucose Isomerase• Previous concepts: Isomerization• Energy use/production? CONCEPT: Near-equilibrium• Chemical logic?• Stereochemistry—reverse does not produce mannose!
3. PFK-1• Previous concepts: Allosteric inhibition• Energy use/production? • Chemical logic?• First committed step of glycolysis
– Why?– regulation
Regulation
4. Aldolase• Previous concepts: Standard free energy is +23kJ, but it
is a near equilibrium reaction• Energy use/production? • Chemical logic?• Beginning of triose stage
Aldolase Mechanism
5. Triose Phosphate Isomerase• Previous concepts: Catalytic perfection• Energy use/production? • Chemical logic?• Most similar to which previous reaction?
6. Glyceraldehyde-3-P DH• Previous concepts: Redox and
dehydrogenase• Energy use/production? • Chemical logic?
GAPDH Mechanism
7. Phosphoglycerate Kinase • Previous concepts: High energy bond• Energy use/production?
– Substrate level phosphorylation• Chemical logic?• Coupled to reaction 6
Coupled Reactions
• GAPDH = 6.7 kJ/mol• PG Kinase = -18.8 kJ/mol• Overall:
8. Phosphoglycerate Mutase• Previous concepts: Covalent catalysis• Energy use/production? • Chemical logic?• Mutase—isomerization with P transfer
Mechanism
• Not a simple transfer• What happens if the bisphosphate escapes?
9. Enolase• Concept: Phosphoryl group transfer potential• Energy use/production? • Chemical logic?
10. Pyruvate Kinase• Energy use/production? • Chemical logic?• Regulation: F-1,6-BP can act as a feed-
forward activator to ensure fast glycolysis
Overall Energetics
• Standard Free energies are up and down
• Free energies under cellular conditions are downhill – Three irreversible
Fate of Pyruvate
Aerobic Energy
Anaerobic inmicroorganisms
Anaerobic inhigher organisms
Gluconeogenesis
Amino acidand nitrogenmetabolism
The Problem of Anaerobic Metabolism
• With oxygen, the NADH produced in glycolysis is re-oxidized back to NAD+
• NAD+/NADH is a co-substrate which means…• If there is no oxygen, glycolysis will stop
because…• The solution to the problem is to…
The solution in Yeast• Pyruvate is decarboxylated
(cofactor?) to acetaldehyde• Acetaldehyde transformed to
ethanol – What type of reaction?– What cofactor?
• NAD+ is regenerated to be reused in GAPDH
The Solution in Us
• Lactate formation
• Balanced equation
We don’t operate anaerobically...
• Most energy still trapped in lactate
• Back to pyruvate, then acetyl-CoA
• Citric acid cycle
Other sugars enter glycolysis
High fructose diet puts sugars through glycolysis while avoiding major regulation step
Glucose Metabolism Overview
• Keep the main pathway purposes distinct
• But learn details of chemistry and regulation based on similarities
O
HO
HO
OH (P)
OH
OH
DHAP
Pyruvate
Gluconeogenesis
Lactate
Amino Acids
Glycerol(Triacylglycerides)
Glycogen
Glycogen Degradation
Glycogen
Glycogen Synthesis
Ribose,NADPH
ATP
DHAP
Pyruvate
Pentose Phosphate Pathway
Energy Production
Starch
Diet
Glucose Metabolism Overview
• Gluconeogenesis• Glycogen
metabolism• Pentose
Phosphate Pathway
O
HO
HO
OH (P)
OH
OH
DHAP
Pyruvate
Gluconeogenesis
Lactate
Amino Acids
Glycerol(Triacylglycerides)
Glycogen
Glycogen Degradation
Glycogen
Glycogen Synthesis
Ribose,NADPH
ATP
DHAP
Pyruvate
Pentose Phosphate Pathway
Energy Production
Precursors for Gluconeogenesis
• Names of compounds?
• Type of reaction?• Type of enzyme?• Cofactor(s)?• More on lactate
processing later…
OH
OH
OH
OPO3
O
OH
O
O
O
NH2
O
O
O
O
O
OH
O
O
Chemistry of Gluconeogenesis
• Chemically opposite of glycolysis (mainly)• Energetically costly—no perpetual motion
machine!• Points of regulation
Glycolysis• Step 1: costs 1 ATP• Step 3: costs 1 ATP• Step 7: makes 2 ATP• Step 10: makes 2
ATP
• Gluconeogenesis• Step 10: no change• Step 8: no change• Step 3: costs 2 ATP• Step 1: costs 4 ATP
equivalents
Step 1a
• Pyruvate Carboxylase– Biotin– Costs ATP to make driving force for next reaction– First step in biosynthesis of glucose and many
other molecules• Related to which amino acid?
Mechanism
• Mixed anhydride• Coupled through
biotin coenzyme
Step 1b
• PEP carboxykinase– ATP cost to restore PEP– CO2 loss drives rxn
Step 8• Fructose-1,6-bisphosphatase• No additional energy input• Phosphate ester hydrolysis is spontaneous
Step 10
• Glucose 6-phosphatase– Liver (and others)– Not in muscle
Problem 34
• A liver biopsy of a four-year old boy indicated that the F-1,6-Bpase enzyme activity was 20% normal. The patient’s blood glucose levels were normal at the beginning of a fast, but then decreased suddenly. Pyruvate and alanine concentrations were also elevated, as was the glyceraldehyde/DHAP ratio. Explain the reason for these symptoms.
Key Regulation• At the committed step in glucogenic cells• Principle of Reciprocal regulation• Local regulation vs Hormone regulation
Key Regulation
• Local regulation– AMP/ATP (energy charge)– Citrate (feedback)
• Hormone regulation– Fructose-2,6-bisphosphate
• Gluconeogenesis is inhibited• Glycolysis is stimulated
Problem 39
• Brazilin, a compound found in aqueous extracts of sappan wood, has been used to treat diabetics in Korea. It increases the activity of the enzyme that products F-2,6-BP and stimulates the activity of pyruvate kinase. What is the effect of adding brazilin to liver cells in culture? Why would brazilin be an effective treatment for diabetes?
Glucose Metabolism Overview
• Gluconeogenesis• Glycogen
metabolism• Pentose
Phosphate Pathway
O
HO
HO
OH (P)
OH
OH
DHAP
Pyruvate
Gluconeogenesis
Lactate
Amino Acids
Glycerol(Triacylglycerides)
Glycogen
Glycogen Degradation
Glycogen
Glycogen Synthesis
Ribose,NADPH
ATP
DHAP
Pyruvate
Pentose Phosphate Pathway
Energy Production
Glycogen
• Storage molecule• Primer necessary• Very large!• Multiple ends allow
for quick synthesis and degradation
Chemistry of Synthesis
• Step 1
• Near equilibrium• The link to glucose-6-phophate, our central
molecule
Chemistry of Synthesis• Step 2• Count high energy
bonds• Pyrophosphatase
– Common motiff• UDP-glucose:
activated for incorporation
Chemistry of Synthesis• Step 3• Glycogen
synthase• Growing end is
non-reducing• UDP released
Energetics of Synthesis
• Total cost: one ATP equivalent from G-6-p
O
HO
HO
O
OH
OH
P
O
O O
O
HO
HO
O
OH
OH
P-P-Uridine
O
HO
OH
OH
OH
O
O
OH
HO
O
HO
Glucose-6-P
UDP
UTP
2 Pi
Chemistry of Degradation
• Glycogen phosphorylase
• Key Regulation site• Inorganic phosphate
as a nucleophile• Remake G-1-P with
no ATP cost
Overall Energetics
O
HO
HO
O
OH
OH
P
O
O O
O
HO
HO
O
OH
OH
P-P-Uridine
O
HO
OH
OH
OH
O
O
OH
HO
O
HO
Glucose-6-P
UDP
UTP
2 Pi
Pi
Key Enzymes
O
HO
HO
O
OH
OH
P
O
O O
O
HO
HO
O
OH
OH
P-P-Uridine
O
HO
OH
OH
OH
O
O
OH
HO
O
HO
Glucose-6-P
UDP
UTP
2 Pi
Pi
Glycogen Synthase
Glycogen Phosphorylase
Glycogen Storage Diseases
Many disrupt glycogen breakdown in muscle and/or liver (hypoglycemia, enlarged liver, muscle cramps...)
Glucose Metabolism Overview
• Gluconeogenesis• Glycogen
metabolism• Pentose
Phosphate Pathway
O
HO
HO
OH (P)
OH
OH
DHAP
Pyruvate
Gluconeogenesis
Lactate
Amino Acids
Glycerol(Triacylglycerides)
Glycogen
Glycogen Degradation
Glycogen
Glycogen Synthesis
Ribose,NADPH
ATP
DHAP
Pyruvate
Pentose Phosphate Pathway
Energy Production
Pentose Phosphate Pathway
• Dual Purpose– Synthesis of “reducing potential”– Synthesis of 5-carbon sugars
• At cost of one carbon worth of carbohydrate• Net reaction:
Complex, 2-Stage Process
• Oxidative Stage– Generates reducing
power and ribose• Non-oxidative stage
– Regenerates 3- and 6-carbon sugars from 5 carbon sugars
Oxidative Stage Step 1:
• G-6-P DH• Lactone formation
Oxidative Stage Step 2:
• Also a spontaneous hydrolysis• Practice mechanism, carbohydrate orientation
Oxidative Stage Step 3:
• Oxidative decarboxylation• We will see this process again
Biosynthesis of Ribose
Non-oxidative Stage• To understand purpose, realize that we
generally need to make much more NADPH than ribose
• Problem: stuck with C5, but need C6 and C3• Solution: “Shunt” C5 back to C6 through near-
equilibrium reactions
PPP Reactions
• Epimerase• Isomerase• Transketolase• Transaldolase
Transketolase• Use cofactor (B1) to overcome chemical problem
Mechanism
Different Modes for Different Purposes
Problem 58
• A given metabolite may follow more than one metabolic pathway. List all possible fates of glucose-6-P in (a) a liver cell and (b) a muscle cell.
Summaryof glucosemetabolism